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ORIGINAL PAPER
Human lysosomal -D-mannosidase regulation in promyelocytic leukaemia cells Lorena Urbanelli, Alessandro Magini, Luisa Ercolani, Francesco Trivelli, Alice Polchi, Brunella Tancini, Carla Emiliani Bioscience Reports Dec 01, 2011, 31 (6) ; DOI: 10.1042/BSR20110020
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Abstract Lysosomal - D-mannosidase is an exoglycosidase involved in the ordered degradation of N-linked oligosaccharides. It is ubiquitously expressed, although the main transcript is more abundant in peripheral blood
December 2011 Volume: 31 Issue: 6
leucocytes. Here we report that - D-mannosidase enzyme activity is very Table of Contents
high in the promyelocytic leukaemia cell lines HL60 and NB4, as compared
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with other leukaemic cell lines or cells from different human sources. The MAN2B1 transcript level correlates with enzyme activity, indicating a transcriptional up-regulation of the - D-mannosidase gene. The promoter was then characterized in HEK-293 cells (human embryonic kidney 293 cells) and HL60 cells; regulatory sequences crucial for its activity were determined by reporter gene assay in HEK-293 cells and located in the region −101/−71 with respect to the first ATG codon. Supershift assay
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demonstrated that Sp1 (specificity protein 1) bound to this sequence both in HEK-293 and HL60 cells. However, 5¢-RACE (5¢-rapid amplification of cDNA ends) indicated the use of multiple upstream TSSs (transcription start sites)
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in HL60 with respect to HEK-293 cells and gel shift analysis of the sequence
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−373/−269 demonstrated a specific binding by NF-B (nuclear factor B)
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transcription factor in HL60 but not in HEK-293 cells. We concluded that
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despite the - D-mannosidase promoter showing typical features of
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housekeeping gene promoters, - D-mannosidase transcription is specifically
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regulated in HL60 by NF-B transcription factor. Abbreviations: AML, acute myeloid leukaemia; C/EBP, CCAAT/enhancerbinding protein; EMSA, electrophoretic mobility-shift assay; HEK, human embryonic kidney; HUDE, human dermal fibroblast(s); NCBI, National
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Center for Biotechnology Information; NE, nuclear extract; NF-B, nuclear factor B; Q-PCR, quantitative PCR; 5¢-RACE, 5¢-rapid amplification of cDNA ends; RQ, relative quantity; Sp1, specificity protein 1; TSS, transcription start site; UTR, untranslated region
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glycohydrolases, Leukaemia, lysosomal -D-
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mannosidase, nuclear factor B (NF-B) , promyelocytic, specificity protein 1 (Sp1)
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